利用重组噬菌体诱导表达的大肠杆菌表达系统

A T7-promoter Based Escherichia coli Expression System Induced with Bacteriophage M13HEP

将编码噬菌体T7RNA聚合酶的基因克隆至噬菌体M13mp18RF DNA中,置于lac启动子的控制之下,得到了可表达T7RNA聚合酶的重组噬菌体M13HEP。利用该噬菌体感染含T7启动子表达质粒的宿主菌以提供T7RNA聚合酶,可以诱导T7启动子控制下的外源基因的表达。该噬菌体诱导表达系统已成功地表达了多种外源基因,特别是一些表达产物对宿主菌有毒性的基因。同时,通过细菌接合将F＇因子从大肠杆菌XL1-blue转至大肠杆菌HMS174,构建了新的大肠杆菌菌株HMS174F＇,使得T7表达质粒构建、表达及单链制备可以在同一菌株中完成,得到了一个完整的T7表达系统。

The bacteriophage M13HEP which can express T7 RNA polymerase under the control of lac promoter was constructed by cloning the T7 RNA polymerase gene into phage M13mp18 RF DNA. Heterologus genes under the control of T7 promoter can be induced by introducing T7 RNA polymerase to the cells through M13HEP phage infection. Many heterologus genes were successfully expressed by using this phage M13HEP induction system, especially some gene products to be expressed are toxic to host strain BL21 (DE3). A new E. coll strain HMS174F' was also constructed by transfer F'pilli from E. coli XL1-blue to E. colt HMS174. It made T7 expression plasmid construction , expression and single strand DNA rescue can be performed in the same E. coli strain.